The overall goal of this project is to evaluate mechanisms that preserve intracellular pH and ion composition in acid-secreting gastric oxyntic cells, under resting conditions and during stimulation of acid secretion. The mechanisms mediating secretion of H+ and Cl ions across the apical membranes of the oxyntic cell are well characterized. Our preliminary studies in in vitro amphibian gastric mucosa suggest that, under some experimental conditions, basolateral Na-K-Cl cotransport plays a dominant and previously unsuspected role in regulating secretion of HCl across the apical membrane. The studies proposed in this application will utilize intracellular microelectrode techniques to evaluate the role of basolateral Na-K-Cl cotransport in regulating intracellular Cl activity (aiCl) and intracellular pH (pHi) in oxyntic cells of Necturus gastric fundus, under resting conditions and during stimulation of acid secretion. The specific aims include studies: 1) to evaluate, under resting conditions and during secretory stimulation, changes in aiCl in oxyntic cells during changes in nutrient ion composition (Cl, Na+, K+) and during exposure to known blockers of Na-K-Cl cotransport; 2) to evaluate the relationship between basolateral Na--k-Cl cotransport and pHi regulation in the oxyntic cell and 3) to evaluate the effects of conditions mimicking metabolic acidosis and respiratory acidosis on regulation of aiCl in oxyntic cells, under resting conditions and during stimulation of acid secretion.. These studies should provide insight into the mechanisms that preserve intracellular pH and ion composition in oxyntic cells during high rates of acid secretion. They may also lead to better understanding of how low perfusion states and respiratory insufficiency predispose gastric mucosa to acute injury.